Project Summary/Abstract The lifespan of individuals living with HIV-1 has significantly increased due to effective combination anti- retroviral therapy. However, new complex pathologies, or HIV-associated non-AIDS conditions are emerging, some of them related to an accelerated or premature aging of the HIV-1-infected population. Chronic infection with HIV-1, persistent immune activation and long-term usage of drugs can all be associated with this premature aging, making patients more at risk for old age cancers and multiple organ disorders. Cellular senescence is an important anti-tumor mechanism implemented by cells in response to a variety of stimuli such as telomere attrition, oncogenic gene activation, and oxidative damage. It is usually characterized by permanent growth arrest, but a variety of phenotypic markers have been used to identify senescent cells such as abnormal cellular enlargement, increased expression of cell cycle regulatory proteins p16(INK4a), p21, and/or p53, presence of senescence-associated heterochromatin foci (SAHF), and a senescence-associated secretory phenotype (or SASP), composed of inflammatory cytokines, chemokines, growth factors, and matrix metalloproteases. Cellular senescence is primarily a beneficial cellular program to prevent tumor growth, but it also promotes cancer and age-related disorders later in life, with senescent cells accumulating in aged tissues, and evidence linking senescent cells with age-related pathologies. Immunosenescence has been well studied but mainly in T and B lymphocytes, and despite some functional alterations in macrophages during aging or HIV-1 infection, macrophage senescence has not been described. Even the concept of ?senescence-after-differentiation?, in terminally-differentiated, non-dividing cells such as neurons or macrophages, is relatively new. Caveolin-1 (Cav- 1), an integral protein component of caveolae, plays a role during oxidative stress and age-related diseases by inhibiting Sirtuin 1 (Sirt1, a protein deacetylase that regulates cell survival and senescence, energy metabolism, inflammation and cancer), leading to acetylation of p53, IL-6 production (a key SASP cytokine), and premature senescence in fibroblasts. In addition, Cav-1 seems to be up-regulated after HIV-1 infection. Our preliminary studies have shown that proteasome inhibition, oxidative damage, continuous culture and HIV-1 infection induce cellular enlargement and Cav-1, p53 and p21 expression, and a reduction in Sirt1 in continuous culture and HIV- 1 infection, suggesting the generation of a non-replicative senescence phenotype in macrophages. Thus, we hypothesize that human macrophages develop a non-replicative senescence phenotype in the context of senescence-associated stimuli and HIV-1 infection, and that this is related with their functional alterations. This hypothesis will be tested by determining: (i) the expression of senescence biomarkers and functional alterations (expression of activation markers, phagocytosis, Toll-like receptor-mediated responses, and cell mediated immunity) in macrophages exposed to senescence stimuli and upon HIV-1 infection; and (ii) the roles of Cav-1 and Sirt1 in the senescence phenotype and associated functional alterations in human macrophages.